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Waterborne Pathogen Prevention and Detection Using Traditional Methods and Microarray Probe Detection Claribel Orellana CE 421 12/5/07 Biotechnology • Priorities: Agriculture and Medicine • • • • • Malaria Vaccine Stem cell research Completion of the human genome map ‘Golden Rice’ modified to make vitamin A GCSF for increasing white blood count in chemotherapy patients World Concern: Water Quality • • • Half the world affected by contaminated water Unsanitary conditions and lack of resources Cause: waterborne pathogens Waterborne Pathogens • Three different types: bacteria, viruses, and bacteria Name of micro-organisms Major diseases Major reservoirs and primary sources Salmonella typhi Typhoid fever Human feces Salmonella paratyphi Paratyphoid fever Human feces Vibrio cholera Cholera Human feces and freshwater zooplankton Enteropathogenic E. coli Gastroenteritis Human feces Yersinia enterocolitica Gastroenteritis Human and animal feces Legionella pneumophila and related bacteria Acute respiratory illness (legionellosis) Thermally enriched water Giardiasis (gastroenteritis) Water and animal feces ascariosis Animal and human feces Bacteria Protozoa Giardia lamblia Helminths Ascaris lumbricoides Typical Contamination • • • • Unprotected water source Inadequate sanitation Animal and fecal matter reaching water source Surface runoff through the ground, water pipes and wells Well Cattle pond Well Surface drain Cattle pond Typical Contamination • • Animal and fecal matter reaching water source Surface runoff through the ground, water pipes and wells Traditional Practices for Safe Water • Point-of-use disinfection • Use of sodium and calcium hypochlorite • Safer and easier to use and distribute • Destroys most pathogens • Electrolysis • System run on solar power • Generators can generate enough disinfectant for 10,000 people. • Inexpensive Traditional Practices for Safe Water • Safe Storage • • Stored water vs. municipal tap Container comparison • CDC container • Cantero DNA Microarrays • • • DNA microarrays: reverse dot-blots for which sequencespecific “probes” are attached to substrate in a lattice pattern spots are usually 100-200 micrometers and 200-500 micrometers away from each other and they represent specific probe sequences Simultaneous detection vs. cultivation Polymerase Chain Reaction (PRC) • PCR amplifies DNA sequences Microarray Process • • Sequences are hybridized Specific bacterial targets are detected Precision Factors • Assay sensitivity • Sample size • Efficiency of pathogen isolation • Efficiency of nucleic acid extraction • Effect of co-precipitating factors that inhibit PCR Benefits and Limitations • Benefits: • • • Simultaneously detects pathogens Not limited to identification by product length Limitations: • • • Requires pathogens to be identified before configuring array Needs to be validated Currently more expensive than traditional methods Microarray Flow Process Application • EPA looking into feasibility • Is being monitored • Will most likely become a standard • Soon be more cost-effective • Priority is quick pathogenic detection
